RNA editing alters pre-mRNA through site-selective adenosine dearnination, which results in codon changes that lead to the production of novel proteins. An enzyme that catalyzes this reaction, double-stranded RNA adenosine deaminase (ADARI), contains two N terminal Z-DNA-binding motifs, Z,,, and 4, the function of which is as yet unknown. In this study, multidimensional NMR spectroscopy was used to show that the topology of Z"' is (X 10 1 (X2 (X3 P2 03. Long-range NOEs indicate that P1 and P3 interact with each other. Site-directed mutagenesis was used to identify residues in 0, 03 and the loop connecting 02 to 03 that affect Z-DNA binding. Also identified were I I hydrophobic residues that are essential for protein stability. Comparison with known structures reveals some similarity between Z,,, and (oc + 0) helix-tum-helix proteins, such as histone 5 and the family of hepatocyte nuclear factor-3 winged-helix-tum-helix transcription factors. Taken together, the structural and functional data suggest that recognition of Z-DNA by Z,,, involves residues in both the 0 helix and the C-terminal P-sheet.